Fig. 6. 3MA reversed FGF21’s effect on random-pattern skin flap viability.

On the 7th day after operation of random flap, the rats were sacrificed in the FGF21 group and FGF21 + 3MA group, and then autophagy level, survival area and subcutaneous microcirculation in the flap were evaluated. a Immunofluorescence for the evaluation of LC3II in the ischemic skin flap (scan bar, 15 μm). b Quantification of percentage of positive cells with LC3II labeled autophagosomes in dermal layer. c, d Western blotting performed for levels of autophagic proteins (Belin1, LC3II, CTSD, VPS34, and p62), which was corrected by GAPDH as internal control. a Histogram showing the quantification of autophagic proteins (Belin1, LC3II, CTSD, VPS34, and p62) detected by western blotting. e Digital photograph of flap survival/necrosis area on the 3rd and 7th day after surgery. f Histogram showing the percentage of survival area of flap on the 7th day. g Digital photograph exhibiting tissue necrosis, edema and the subcutaneous vascular network on the inner surface of the flap. h Histogram showing the percentage of tissue water content of flap. i The subcutaneous vascular network detected by LDBF, with stronger signal intensity representing greater blood flow. j Histogram showing signal intensity of blood flow in flap. k H&E staining exhibiting subcutaneous histology of the flap, showing subcutaneous blood vessels and inflammation (original magnification × 200; scan bar, 50 μm). l Histogram showing mean vessel density (MVD) in the flap (/mm²). m IHC of CD34 which mainly were expressed in vascular endothelial cells. n Histogram showing the density of CD34-positive blood vessels (/mm²). Significance: *p < 0.05 and **p < 0.01 vs the FGF21 group. Data were expressed as means ± SEM (n = 6 per group).